Received for publication May 15, 2007.
Revised September 15, 2007.
Accepted for publication September 17, 2007.

QUANTIFICATION OF pCREB EXPRESSION THROUGHOUT THE BRAIN OF AMPHETAMINE SENSITIZED RATS: ACTIVATION OF HYPOTHALAMIC OREXIN A-CONTAINING NEURONS

Abstract

In the present study using rats, we have examined acute, contextual and sensitized patterns of activated or phosphorylated CREB (pCREB) expression in parallel, assaying across multiple nuclei which have been implicated in addiction. The paradigm employed included a comparison of pre-treatment dose of amphetamine upon patterns of cellular activation, following re-challenge. As efferent orexinergic projections synapse on many targets through the mammalian brain, including mesotelencephalic regions and limbic systems involved in drug reward and reinforcement, we examined for co-expression of pCREB or c-Fos double-labelling within orexin A-immunopositive neurons following sensitization. Acute challenge with amphetamine (1.5 mg/kg, i.p.) resulted in an increase in the number of pCREB-immunoreactive (-IR) cells within the substantia nigra but a decrease of pCREB-IR cells in the central and medial subnuclei of the amygdala. Contextual re-exposure to the drug treatment environment altered pCREB expression, particularly in the basal ganglia and hypothalamus, although these effects were dictated by pre-treatment dose of amphetamine. Sensitization to amphetamine resulted in robust increases in pCREB-IR cell numbers in the basal ganglia and lateral septum of rats that had been pre-treated with high dose (10 mg/kg, i.p.) but not low dose (2 mg/kg, i.p.) amphetamine, despite a similar behavioral response. Orexin A-containing cells in the hypothalamus of sensitized rats did not co-express pCREB; however, these cells double-labelled for c-Fos and orexin A. These data suggest that orexinergic neurons are activated during the expression of behavioral sensitization, although in a heterogenous manner with regards to afferent topologies and functional roles in the nervous system.

Key words: Amphetamine, c-Fos, immunohistochemistry, orexin, pCREB, sensitization